The long term objective of this proposal is to understand how a Gram negative bacterial pathogen, Legionella pneumophila, is able to survive and replicate inside normally bactericidal phagocytic cells. This grant will focus on determining how L. pneumophila alters the endocytic pathway and prevents phagosome-lysosome fusion of host cells. The specific aims are to characterize the Dot apparatus by examining the role of two critical Dot proteins, DotB and DotL, in the assembly and activity of the Dot/Icm complex and to identify the substrate(s) exported by this complex into the macrophage host cell. The health relatedness of this proposal is two fold. First, establishing the mechanism used by L, pneumophila to survive and replicate inside macrophages will provide additional insight into how it causes disease and may reveal novel targets to be used for drug therapy. Second, since specialized secretion systems are commonly used by a variety of bacterial pathogens, knowledge gained about the L. pneumophila secretion apparatus is likely to be applicable to understanding the molecular mechanisms of virulence used by other pathogens, and could serve as the basic to prevent or treat a number of different disease.